view scpipe.R @ 0:32e1bfc6b7b2 draft

planemo upload for repository https://github.com/galaxyproject/tools-iuc/tree/master/tools/scpipe commit 8908da9cdd112ae0943dbf1eccb221e84cd99ca7

options( show.error.messages=F, error = function () { cat( geterrmessage(), file=stderr() ); q( "no", 1, F ) } )

# we need that to not crash galaxy with an UTF8 error on German LC settings.
loc <- Sys.setlocale("LC_MESSAGES", "en_US.UTF-8")

suppressPackageStartupMessages({
    library(scPipe)
    library(SingleCellExperiment)
    library(optparse)
    library(readr)
    library(ggplot2)
    library(plotly)
    library(DT)
    library(scater)
    library(scran)
    library(scales)
    library(Rtsne)
})

option_list <- list(
    make_option(c("-fasta","--fasta"), type="character", help="Genome fasta file"),
    make_option(c("-exons","--exons"), type="character", help="Exon annotation gff3 file"),
    make_option(c("-barcodes","--barcodes"), type="character", help="Cell barcodes csv file"),
    make_option(c("-read1","--read1"), type="character", help="Read 1 fastq.gz"),
    make_option(c("-read2","--read2"), type="character", help="Read 2 fastq.gz"),
    make_option(c("-samplename","--samplename"), type="character", help="Name to use for sample"),
    make_option(c("-bs1","--bs1"), type="integer", help="Barcode start in Read 1"),
    make_option(c("-bl1","--bl1"), type="integer", help="Barcode length in Read 1"),
    make_option(c("-bs2","--bs2"), type="integer", help="Barcode start in Read 2"),
    make_option(c("-bl2","--bl2"), type="integer", help="Barcode length in Read 2"),
    make_option(c("-us","--us"), type="integer", help="UMI start in Read 2"),
    make_option(c("-ul","--ul"), type="integer", help="UMI length in Read 2"),
    make_option(c("-rmlow","--rmlow"), type="logical", help="Remove reads with N in barcode or UMI"),
    make_option(c("-rmN","--rmN"), type="logical", help="Remove reads with low quality"),
    make_option(c("-minq","--minq"), type="integer", help="Minimum read quality"),
    make_option(c("-numbq","--numbq"), type="integer", help="Maximum number of bases below minq"),
    make_option(c("-stnd","--stnd"), type="logical", help="Perform strand-specific mapping"),
    make_option(c("-max_mis","--max_mis"), type="integer", help="Maximum mismatch allowed in barcode"),
    make_option(c("-UMI_cor","--UMI_cor"), type="integer", help="Correct UMI sequence error"),
    make_option(c("-gene_fl","--gene_fl"), type="logical", help="Remove low abundant genes"),
    make_option(c("-max_reads","--max_reads"), type="integer", help="Maximum reads processed"),
    make_option(c("-min_count","--min_count"), type="integer", help="Minimum count to keep"),
    make_option(c("-report","--report"), type="logical", help="HTML report of plots"),
    make_option(c("-rdata","--rdata"), type="logical", help="Output RData file"),
    make_option(c("-nthreads","--nthreads"), type="integer", help="Number of threads")
  )

parser <- OptionParser(usage = "%prog [options] file", option_list=option_list)
args = parse_args(parser)

fa_fn = args$fasta
anno_fn = args$exons
fq_R1 = args$read1
fq_R2 = args$read2
read_structure = list(
    bs1 = args$bs1,   # barcode start position in fq_R1, -1 indicates no barcode
    bl1 = args$bl1,    # barcode length in fq_R1, 0 since no barcode present
    bs2 = args$bs2,    # barcode start position in fq_R2
    bl2 = args$bl2,   # barcode length in fq_R2
    us = args$us,    # UMI start position in fq_R2
    ul = args$ul     # UMI length
)

if (args$us == -1) {
  has_umi = FALSE
} else {
  has_umi = TRUE
}

filter_settings=list(rmlow=args$rmlow, rmN=args$rmN, minq=args$minq, numbq=args$numbq)

# Outputs
out_dir = "."
fasta_index = file.path(out_dir, paste0(fa_fn, ".fasta_index"))
combined_fastq = file.path(out_dir, "combined.fastq")
aligned_bam = file.path(out_dir, "aligned.bam")
mapped_bam = file.path(out_dir, "aligned.mapped.bam")


print("Trimming barcodes")
sc_trim_barcode(combined_fastq,
                fq_R1,
                fq_R2,
                read_structure=read_structure,
                filter_settings=filter_settings)

print("Building genome index")
Rsubread::buildindex(basename=fasta_index, reference=fa_fn)

print("Aligning reads to genome")
Rsubread::align(index=fasta_index,
    readfile1=combined_fastq,
    output_file=aligned_bam,
    nthreads=args$nthreads)

if (!is.null(args$barcodes)) {
  barcode_anno=args$barcodes
} else {
  print("Detecting barcodes")
  # detect 10X barcodes and generate sample_index.csv file
  barcode_anno = "sample_index.csv"
  sc_detect_bc(
      infq=combined_fastq,
      outcsv=barcode_anno,
      bc_len=read_structure$bl2,
      max_reads=args$max_reads,
      min_count=args$min_count,
      max_mismatch=args$max_mis
  )
}

print("Assigning reads to exons")
sc_exon_mapping(aligned_bam, mapped_bam, anno_fn, bc_len=read_structure$bl2, UMI_len=read_structure$ul, stnd=args$stnd)

print("De-multiplexing data")
sc_demultiplex(mapped_bam, out_dir, barcode_anno, has_UMI=has_umi)

print("Counting genes")
sc_gene_counting(out_dir, barcode_anno, UMI_cor=args$UMI_cor, gene_fl=args$gene_fl)

print("Creating SingleCellExperiment object")
sce <- create_sce_by_dir(out_dir)

if (!is.null(args$report)) {
print("Creating report")
create_report(sample_name=args$samplename,
   outdir=out_dir,
   r1=fq_R1,
   r2=fq_R2,
   outfq=combined_fastq,
   read_structure=read_structure,
   filter_settings=filter_settings,
   align_bam=aligned_bam,
   genome_index=fasta_index,
   map_bam=mapped_bam,
   exon_anno=anno_fn,
   stnd=args$stnd,
   fix_chr=FALSE,
   barcode_anno=barcode_anno,
   max_mis=args$max_mis,
   UMI_cor=args$UMI_cor,
   gene_fl=args$gene_fl)
}

if (!is.null(args$rdata) ) {
    save(sce, file = file.path(out_dir,"scPipe_analysis.RData"))
}

sessionInfo()